During the past cycles of this P41, this project has been primarily focused on the development and dissemination of continuous-wave near infrared spectroscopy (CW-NIRS) for functional brain imaging in humans. We have commercialized instrumentation, distributed open source code for analyzing CW-NIRS data that has been downloaded by more than 1000 unique users, ran numerous training workshops, supported a number of service users in varied capacities, and founded the Functional Near Infrared Spectroscopy Society. This technology continues to mature and experience exponential growth, and we will continue to support service users under this P41. However, our new aims for technology research and development for this competitive renewal depart from CW-NIRS. This reflects the growing driving forces arising from our collaborative projects pushing us to develop novel optical methods to address questions that cannot be addressed by CW-NIRS. While CW-NIRS excels at imaging changes in hemoglobin concentrations within the brain, it cannot quantify baseline hemoglobin concentrations (as can be done by Time Domain NIRS), nor does it provide a direct measure of blood flow (as provided by Diffuse Correlation Spectroscopy). The ability to measure these hemodynamic parameters is important for a number of applications including ensuring sufficient oxygen delivery to the brain following brain injury (CP Franceschini), during epileptic seizures (CP Stufflebeam), and during anesthesia (CP Brown). By combining measurements of oxygenated and deoxygenated hemoglobin with measurements of blood flow, we can estimate the cerebral metabolic rate of oxygen (CMRO2), a measure of energy metabolism within the brain. This is important for understanding the severity of brain injury and the response to therapy (CP Franceschini), the metabolic impact of epileptic seizures (CP Stufflebeam), and the depth of anesthesia (CP Brown). In addition, reduced cerebral energy metabolism in the dorso-lateral prefrontal cortex has been observed by PET in depression and shown to normalize during therapy [Mayberg2000, Dougherty2003]. An all-optical method that can be used in an office without the need for radio nucleotides would enable more Wide spread investigation of the pathophysiology of depression and potentially provide an early indicator of treatment efficacy (CP Haber). In parallel with advancing novel methods for non-invasive optical spectroscopy and imaging of humans, our lab has a history of developing optical microscopy methods for studying the brain [Sakadzic2010, Srinivasan2011, Lee2013]. Responding to the significant need of TRD 1 to support collaborative projects (CPs Milad, Haber, Mayberg) in depression and anxiety disorders, we've been utilizing Optical Coherence Tomography (OCT) to provide histology grade images from block face tissue samples. The block face images provide cytoarchitectural boundaries with 10's pm resolution that are easily registered with the MRI images to train segmentation algorithms (TRD 1) to then support these CPs. This overcomes the significant challenges presented by standard histology in registering with MRI for training segmentation algorithms.